Qualitative Analysis of Tissue Biomarkers and Pathways

组织生物标志物和通路的定性分析

• 批准号: 6957728
• 负责人: ROBERT L CAMP
• 金额: $ 14.06万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-07 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6957728
• 关键词: biomarker; clinical research; conformation; crosslink; fluorescence resonance energy transfer; human tissue; immunocytochemistry; neoplastic growth; protein protein interaction; sectioning; technology /technique development

DESCRIPTION (provided by applicant): Quantitative measurement of tumor biomarkers has shown great promise in predicting patient outcome and response to treatment. Recently, we have developed an automated system for assessing biomarker expression on tissue sections (AQUA - Automated Quantitative Analysis). AQUA provides quantitative analysis and sub-cellular localization of biomarkers on immunohistochemically stained tissues. AQUA, however, is limited by 1) the resolution of light microscopy and 2) the non-linear effects of enzymatic amplification used to identify biomarkers. The progression toward bio-specific therapies and associated pharmaco-diagnostics will require linear, quantitative measures of protein expression within each patient's tumor. Individual tumor profiling can also be advanced by developing methods for assessing the functioning of tumor-related pathways, in particular by studying specific protein-protein interactions. To acomplish this, we propose the following specific aims: 1: To develop methods for identifying specific protein-protein interactions in tissue sections. And 2: To develop methods for increasing the dynamic range of assessing biomarker expression. To assess protein-protein interactions - which are far below the resolution of light microscopy - we will develop new techniques that combine heterobifunctional crosslinking reagents, fluorescence energy resonance transfer (FRET), and catalyzed tyramide-amplification, making them suitable for use in tissue sections. To improve the dynamic range of the AQUA technology, we will adapt several recently described methods for building large fluorescent molecules in situ (Christmas trees, rolling-circle and ImmunoAT-tailing). Once these techniques are developed, they will provide new linear, quantitative, and highly specific assessments of biomarkers and pathways in tissues that can ultimately be used to advance personalized pharmaco-diagnostics.

描述（由申请人提供）：肿瘤生物标志物的定量测量在预测患者结果和治疗反应方面显示出巨大的前景。最近，我们开发了一种用于评估组织切片上生物标志物表达的自动化系统（AQUA - 自动定量分析）。 AQUA 提供免疫组织化学染色组织上生物标志物的定量分析和亚细胞定位。然而，AQUA 受到 1) 光学显微镜分辨率和 2) 用于识别生物标志物的酶扩增的非线性效应的限制。生物特异性疗法和相关药物诊断的进展将需要对每个患者肿瘤内的蛋白质表达进行线性、定量测量。个体肿瘤分析也可以通过开发评估肿瘤相关通路功能的方法来推进，特别是通过研究特定的蛋白质-蛋白质相互作用。为了实现这一目标，我们提出以下具体目标： 1：开发识别组织切片中特定蛋白质-蛋白质相互作用的方法。 2：开发增加评估生物标志物表达的动态范围的方法。为了评估远低于光学显微镜分辨率的蛋白质-蛋白质相互作用，我们将开发结合异双功能交联试剂、荧光能量共振转移（FRET）和催化酪酰胺扩增的新技术，使它们适合在组织切片中使用。为了提高 AQUA 技术的动态范围，我们将采用几种最近描述的原位构建大荧光分子的方法（圣诞树、滚环和免疫 AT 拖尾）。一旦开发出这些技术，它们将为组织中的生物标志物和通路提供新的线性、定量和高度特异性的评估，最终可用于推进个性化药物诊断。